丙型肝炎病毒1b型NS5A区基因复杂性与干扰素α疗效的关系

Relationship between the complexity of HCV-1b NS5A genes and the therapeutic response to interferon-α

目的 探讨丙型肝炎病毒 1b型 (HCV 1b)NS5A区基因的复杂性与干扰素α(IFN α)治疗效果之间的关系。方法 以 13例接受IFN α治疗的HCV 1b感染患者为研究对象 ,治疗前血清提取的RNA采用逆转录套式PCR (RT nested PCR)扩增HCVNS5A区基因 ,PCR产物纯化后与T载体连接并克隆入大肠杆菌 ,随机挑选 30个阳性克隆菌落 ,每一克隆菌落再进行PCR扩增 ,30个克隆的PCR产物在同一聚丙烯酰胺凝胶上采用单链构象多态性分析 (SSCP)分析和异源性双体分析 (HD)筛选HCVNS5A区克隆型 ,克隆型的多少反映HCVNS5A区基因的复杂程度。结果 完全应答组平均 6 .3种克隆型 ;部分应答组平均 8.6种克隆型 ;无应答组平均 15 .8种克隆型。结论 HCVNS5A区基因的复杂程度与IFN α的治疗效果呈负相关。

Objective To investigate the relationship between the complexity of HCV 1b NS5A genes and therapeutic response to interferon alpha (IFN α). Methods All of the 13 patients studied were infected with HCV 1b and treated with IFN α, but therapeutic response to IFN α were different. HCV RNA was extracted from pretreatment serum and HCV NS5A gene was amplified by reverse transcription nested PCR. The PCR products were connected to T vector after purification, and cloning. Thirty positive clones were randomly selected to amplify again with PCR. Clonotypes were determined by single strand assay ( single stranded conformational polymorphism, SSCP) and double strand assay (heteroduplex analysis, HD) on the same gel by polyacrylamide electrophoresis (PAGE). The number of clonotypes represented the complexity of HCV NS5A genes. Results The group with complete response, partial response and non response had 6.3 clonotypes, 8.6 clonotypes and 15.8 clonotypes respectively on average. Conclusions There exists a negative correlation between the HCV NS5A gene complexity and the therapeutic efficacy of IFN α.